The goal of this project is to identify novel therapeutic targets for HIV-1 infection, focusing on cellular co- factors involved in the viral Tat protein or Rev protein function. Although it is not known how many cellular factors are involved in Tat and Rev function, recent studies and data mining have revealed that on the order of 50 cellular proteins are likely involved in these processes. The best understood cellular co-factor for Tat is a protein kinase complex termed P-TEFb. P-TEFb is composed of Cyclin T1 (CCNT1) as a regulatory subunit and CDK9 as the catalytic subunit; Tat binds directly to CCNT1. The best understood cellular co-factor for Rev is termed XPO1 (CRM1); Rev binds directly to XPO1. Nearly all processes in cells are carried out by multiprotein complexes which are thought to be the minimal functional unit. It is therefore likely that both CCNT1/CDK9 and XPO1 exist is several higher order multiprotein complexes and these complexes may be involved in Tat and Rev function. We have mined a recently published dataset that described the human nuclear complexome - the set of multiprotein complexes defined by a high-throughput co-immunoprecipitation/mass spec analysis. We have identified a number of novel cellular complexes that contain CCN1/CDK9 or XPO1. We will test the hypothesis that several of these complexes are involved in either Tat or Rev function. In our preliminary studies, we have obtained strong experimental support for this hypothesis. Importantly, our initial results also indicate that we will identify cellular factors that negatively regulate Tat activation of the HIV- LTR. Such negative acting factors have therapeutic potential in strategies to purge the latent HIV-1 reservoir in patients taking anti-viral drugs. Completion of the proposed work will reveal a number of potential therapeutic targets, especially some that may be useful for strategies to purge the reservoir of latent viruses in HIV-infected individuals. PUBLIC HEALTH RELEVANCE: We will identify cellular proteins that regulate HIV-1 replication. Some of these proteins might be useful as therapeutic targets to treat infection. Our research may improve patient treatment.